Cyanidin-3-glucoside inhibits glutamate-induced Zn2+ signaling and neuronal cell death in cultured rat hippocampal neurons by inhibiting Ca2+-induced mitochondrial depolarization and formation of reactive oxygen species

摘要

Cyanidin-3-glucoside (C3G), a member of the anthocyanin family, is a potent natural antioxidant. However, effects of C3G on glutamate-induced [Zn2+] increase and neuronal cell death remain unknown. We studied the effects of C3G on glutamate-induced [Zn2+] increase and cell death in cultured rat hippocampal neurons from embryonic day 17 maternal Sprague-Dawley rats using digital imaging methods for Zn2+, Ca2+, reactive oxygen species (ROS), mitochondrial membrane potential and a MTT assay for cell survival. Treatment with glutamate (100 mu M) for 7 min induces reproducible [Zn2+](i) increase at 35 min interval in cultured rat hippocampal neurons. The intracellular [Zn2+]-chelator TPEN markedly blocked glutamate-induced [Zn2+] increase, but the extracellular Zn2+ chelator CaEDTA did not affect glutamate-induced [Zn2+] increase. C3G inhibited the glutamate-induced [Zn2+] response in a concentration-dependent manner (IC50 of 14.1+/-1.1 mu g/ml). C3G also significantly inhibited glutamate-induced [Ca2+] increase. Two antioxidants such as Trolox and DTT significantly inhibited the glutamate-induced [Zn2+](i) response, but they did not affect the [Ca2+](i) responses. C3G blocked glutamate-induced formation of ROS. Trolox and DTT also inhibited the formation of ROS. C3G significantly inhibited glutamateinduced mitochondrial depolarization. However, TPEN, Trolox and DTT did not affect the mitochondrial depolarization. C3G, Trolox and DTT attenuated glutamate-induced neuronal cell death in cultured rat hippocampal neurons, respectively. Taken together, all these results suggest that cyanidin-3-glucoside inhibits glutamate-induced [Zn2+](i) increase through a release of Zn2+ from intracellular sources in cultured rat hippocampal neurons by inhibiting Ca2+-induced mitochondrial depolarization and formation of ROS, which is involved in neuroprotection against glutamate-induced cell death. (C) 2015 Elsevier B.V. All rights reserved.